Molecular characterization of   from   and   and  -based discrimination of staphylococcal species

Vannuffel, P.; Heusterspreute, Michel; Bouyer, Michele; Vandercam, Bernard; Philippe, Marianne; Gala, Jean‐Luc

doi:10.1016/s0923-2508(99)80030-8

Cited by 56 publications

(42 citation statements)

References 26 publications

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“…It remains an issue whether CoNS molecular identification at the species level is of clinical value (31), but recent findings indicate that some CoNS species, such as S lugdunensis (patient 2), may have virulence factors that closely resemble those of S aureus, and consequently, they may cause similar aggressively destructive infections (32). Regarding typing of staphylococci, it is also interesting that reverse hybridization of biotin-labeled nested femA and mecA amplicons on a prototype staphylococci-and methicillin resistance-specific DNA array yielded results that were fully concordant with those of the sequence analysis performed on 16S rDNA and/or femA amplicons (17,19). Although performed on a very limited number of case samples, this innovative assay indeed opens the way to the development and use of array technology for detection and typing of staphylococci in biologic fluids and tissues.…”

Section: Diagnosing Infectious Discitis By Dna Analysismentioning

confidence: 70%

“…The original methods were modified by the use of a nested consensus PCR rather than a single-round PCR. Briefly, a first PCR was performed with consensus primers for staphylococcal genomic DNA (19), and a second PCR was then performed with nested degenerated consensus primers as described (17) ( Table 1). The amplicons generated by the nested PCR were sequenced, and the results were compared with the femA sequences from GenBank (17,19).…”

Section: Diagnosing Infectious Discitis By Dna Analysis 2987mentioning

confidence: 99%

“…Briefly, a first PCR was performed with consensus primers for staphylococcal genomic DNA (19), and a second PCR was then performed with nested degenerated consensus primers as described (17) ( Table 1). The amplicons generated by the nested PCR were sequenced, and the results were compared with the femA sequences from GenBank (17,19). Determination of the staphylococcal susceptibility to methicil- * PCR ϭ polymerase chain reaction; Gϩ ϭ gram positive; GϪ ϭ gram negative; rDNA ϭ ribosomal DNA; cons-s ϭ consensus sense; F ϭ forward; as ϭ antisense; R ϭ reverse.…”

Section: Diagnosing Infectious Discitis By Dna Analysis 2987mentioning

confidence: 99%

“…Capture probes are covalently attached by an amino group located at the 5Ј end onto the aldehyde-activated slide. The prototype array contains 5 selective DNA capture probes (S aureus, Staphylococcus epidermidis, Staphylococcus haemolyticus, Staphylococcus hominis, and Staphylococcus saprophyticus), a consensus capture probe that detects all femA sequences and allows the identification of the genus Staphylococcus, and a specific capture probe for the mecA gene (19). Duplex femA and mecA amplification was carried out directly on the clinical sample, as described above.…”

Section: Identification Of Staphylococci By Hybridization Of Ampliconmentioning

confidence: 99%

“…Regarding discrepancies between molecular and microbiologic methods, the former method was successful in 5 culture-negative cases (S simulans [ 19]). In the last case, cloning of barely visible amplicons was mandatory to enable sequence analysis, but the molecular result strengthened the clinical and radiographic suspicions of tuberculous discitis.…”

Section: Diagnosing Infectious Discitis By Dna Analysismentioning

confidence: 99%

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The etiologic diagnosis of infectious discitis is improved by amplification‐based DNA analysis

Lecouvet¹,

Irenge²,

Vandercam³

et al. 2004

Arthritis & Rheumatism

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Objective. Blood cultures and cultures of disc material are required to identify and treat bacterial agents responsible for septic spondylodiscitis, but these methods have limited sensitivities. We undertook this study to compare nonculture amplification-based DNA analysis with conventional culture of disc aspirate.Methods. Nineteen patients with spondylodiscitis, including 11 with a history of spinal surgery, presented with negative blood cultures and underwent percutaneous disc or epidural abscess puncture for bacterial diagnosis. Amplification by polymerase chain reaction was performed on 16S ribosomal DNA universal target genes and femA staphylococci-specific target genes in all patients, and on the upstream p34 mycobacterial gene in 1 patient. Species identification relied on amplicon sequencing and comparison with templates from GenBank. Amplification of the femA gene led to subsequent testing for methicillin resistance by amplification of the mecA gene. Further assessment using a staphylococciand methicillin resistance-specific DNA array was performed on 3 samples.Results. Microbiologic and molecular assays identified the causative organism in 14 of 19 patients (74%) and 19 of 19 patients (100%), respectively. In culture-positive patients, DNA-based and microbiologic results were highly correlated. Five agents (Staphylococcus simulans, Staphylococcus sciuri, Brucella species, Actinomyces israelii, and Mycobacterium tuberculosis complex) were identified only by DNA-based methods. In 1 sample, Corynebacterium jeikeium and coagulasenegative Staphylococcus were both cultured, whereas DNA analysis identified only Staphylococcus hominis.Conclusion. DNA-based methods are highly sensitive and specific. They can usefully complement standard microbiologic methods for identifying the cause of infectious spondylodiscitis and contribute to speciesspecific therapeutic orientation in patients with negative blood and disc aspirate cultures.Infectious spondylodiscitis consists of concurrent infection of the intervertebral disc and adjacent vertebral osteomyelitis, observed spontaneously or after invasive spinal procedures or surgery (1-3). Magnetic resonance imaging (MRI) is the diagnostic technique of choice, with sensitivity and specificity of 96% and 94%, respectively, whereas radiographic abnormalities only appear several weeks or months after clinical symptoms (4,5).Given the wide spectrum of potential pathogens (Staphylococcus aureus, coagulase-negative staphylococci [CoNS], streptococci, gram-negative bacilli, Mycobacterium species, fungi), definite identification of the microbial agent is mandatory before initiation of longduration and appropriate parenteral antibiotic therapy. In the absence of bacteremia, percutaneous disc aspiration or biopsy is performed for pathogen identification, but conventional microbiologic techniques fail to identify the causative agent in 30-50% of patients (2,6-9).Nonculture molecular methods based on the amplification and/or sequence analysis of conserved or species-specific genomic ...

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Section: Diagnosing Infectious Discitis By Dna Analysismentioning

confidence: 70%

Section: Diagnosing Infectious Discitis By Dna Analysis 2987mentioning

confidence: 99%

Section: Diagnosing Infectious Discitis By Dna Analysis 2987mentioning

confidence: 99%

Section: Identification Of Staphylococci By Hybridization Of Ampliconmentioning

confidence: 99%

Section: Diagnosing Infectious Discitis By Dna Analysismentioning

confidence: 99%

See 3 more Smart Citations

The etiologic diagnosis of infectious discitis is improved by amplification‐based DNA analysis

Lecouvet¹,

Irenge²,

Vandercam³

et al. 2004

Arthritis & Rheumatism

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Untitled

2009

EFSA Journal

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SUMMARYThe European Food Safety Authority (EFSA) asked its Panel on Biological Hazards to deliver a scientific opinion on: The assessment of the Public Health significance of meticillin resistant Staphylococcus aureus (MRSA).There are different states of interaction between S. aureus and its host: infections, carriage or colonisation, and contamination. Meticillin resistant S. aureus (MRSA) can be persistently or intermittently carried by healthy humans, and colonisation is the major risk factor for infection. Infection can be mild to severe and, in some instances, fatal. MRSA are now a major cause of hospital acquired infection in many European countries, with large differences in prevalence and control policies. A limited number of lineages of MRSA tend to predominate in specific geographical locations. CC398 is the MRSA lineage most often associated with asymptomatic carriage in intensively reared food-producing animals. MRSA commonly carry enterotoxin genes but there has been only one report of food intoxication due to MRSA. On the question on what is the risk to human health posed by MRSA associated with foodproducing animals, the Panel concluded that:Livestock-associated MRSA (LA-MRSA) represent only a small proportion of the total number of reports of MRSA infections in the EU. However, this proportion differs between Member States. In some countries with low prevalence of human MRSA infection, CC398 is a major contributor to the overall MRSA burden. In countries with high overall human MRSA prevalence, CC398 is considered of less significance for the public health. CC398 has, albeit rarely, been associated with deep-seated infections of skin and soft tissue, pneumonia and septicaemia in humans. Where CC398 prevalence is high in food-producing animals, people in contact with these live animals (especially farmers and veterinarians, and their families) are at greater risk of colonisation and infection than the general population. The risk to human health from different levels (dose response) of MRSA during carriage in animals (and in the environment) is not known. On the question of what is the importance of food, food-producing animals, and companion animals in the risk of human infection and/or food-borne disease caused by MRSA in both the community and hospital settings, the Panel concluded that:Food may be contaminated by MRSA (including CC398): eating and handling contaminated food is a potential vehicle for transmission. There is currently no evidence for increased risk of human colonisation or infection following contact or consumption of food contaminated by CC398 both in the community and in hospital. MRSA (including CC398) can enter the slaughterhouse in or on animals and occurs on raw meat. Although it may become part of the endemic flora of the slaughterhouse, the risk of infection to slaughterhouse workers and persons handling meat appears to be low based on currently available data.Where CC398 prevalence is high in food-producing animals, people in direct contact with these live animals (espe...

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